The expression of four major steroidogenic enzymes in porcine theca and granulosa cell layers of preovulatory follicles was related to the levels of steroids in follicular fluid and gonadotropin concentrations in peripheral serum at slaughter. Ovaries were collected during proestrus, early estrus, and late estrus as evidenced by behavioral signs. Follicles were dissected from the ovaries, and theca, granulosa, and follicular fluids were pooled for each of 24 sows. Cytochromes P450 17 alpha-hydroxylase/17-20 lyase (P450c17), aromatase (P450arom) and side-chain cleavage (P450scc), as well as 3 beta-hydroxysteroid dehydrogenase (3 beta HSD), were subjected to Northern and Western immunoblot analyses. The concentrations of estradiol-17 beta, testosterone, androstenedione, and progesterone were determined in follicular fluid, and peripheral serum was assayed for estradiol-17 beta, LH, and FSH. Stages of preovulatory development were verified by plasma levels of LH, FSH, and estradiol-17 beta. Thecca expressed P450c17, P450arom, P450scc, and 3 beta HSD whereas granulosa expressed only P450arom and low levels of P450scc. Thecal P450c17, thecal P450arom, and granulosa P450arom expression decreased coincidentally as serum estradiol-17 beta and follicular fluid estradiol-17 beta, testosterone, and androstenedione levels declined after the presumed gonadotropin surge. Unlike P450c17 and P450arom P450scc and 3 beta HSD remained relatively constant in theca and granulosa. From these data, we suggest that the theca interna may be the primary steroidogenic compartment of the porcine follicle during its final stages of preovulatory development. Moreover, preovulatory estrogen secretion appears to be controlled by the coordinated expression of a triad of enzymes in the porcine follicle that includes theca P450c17, theca P450arom and granulosa P450arom.